Beijing Northland Biotech.Co., Ltd.

Therapeutic angiogenesis offers a promising strategy for patients with critical limb-threatening ischemia (CLTI) who are unsuitable candidates for revascularization. However, the optimal administration sites for gene therapy agents, such as pCK-HGF-X7, remains undefined. Clinical trials commonly employ multiple intramuscular injections at sites of arterial occlusion; yet the necessity and efficacy of such extensive and repetitive protocols remains unclear. Targeted injections into ischemic tissues or their margins may improve therapeutic outcomes. Moreover, the molecular mechanisms by which hepatocyte growth factor (HGF)/c-Met signaling regulates hypoxia-inducible factor-1α (HIF-1α) expression under hypoxic conditions are not fully understood. This study aims to elucidate these molecular mechanisms in endothelial cells under hypoxic conditions and to identify the most effective injection sites for therapeutic angiogenesis agents. The effects of various HGF isoforms/complexes on human aortic endothelial cells (HAECs) were evaluated under normoxic and hypoxic conditions, focusing on proliferation, migration, and tube formation. Pathway inhibitors were used to explore the underlying mechanisms in hypoxic HAECs, and the findings were validated in a rat hindlimb ischemia model. Results demonstrated that HGF₇₂₃ and HGF₇₂₈ activated the mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K) pathways, regulating HIF expression and significantly enhanced endothelial cell proliferation, migration, and tube formation, particularly under hypoxia. Despite these cellular effects, HGF treatment did not significantly improve tissue perfusion or neovascularization in normal rat hindlimbs. However, in ischemic rat hindlimbs, it markedly promoted angiogenesis and improved tissue perfusion in the gastrocnemius muscle. These findings indicate that therapeutic angiogenesis agents should primarily target hypoxic tissues, extending to the interface between normoxic and hypoxic regions, to optimize treatment efficacy.

We investigated the therapeutic effect of recombinant thymosin β4 (rTβ4) on rabbit autoimmune dacryoadenitis, an animal model of SS dry eye, and explore its mechanisms.

Rabbits were treated topically with rTβ4 or PBS solution after disease onset for 28 days, and clinical scores were determined by assessing tear secretion, break-up time, fluorescein, hematoxylin and eosin staining, and periodic acid-Schiff. The expression of inflammatory mediators in the lacrimal glands were measured by real-time PCR. The expression of T helper 17 (Th17) cell-related transcription factors and cytokines were detected by real-time PCR and Western blotting. The molecular mechanism underlying the effects of rTβ4 on Th17 cell responses was investigated by Western blotting.

Topical administration of rTβ4 after disease onset efficiently ameliorated the ocular surface inflammation and relieved the clinical symptoms. Further analysis revealed that rTβ4 treatment significantly inhibited the expression of Th17-related genes (RORC, IL-17A, IL-17F, IL-1R1, IL-23R, and granulocyte-macrophage colony-stimulating factor) and IL-17 protein in lacrimal glands, and meanwhile decreased the inflammatory mediators expression. Mechanistically, we demonstrated that rTβ4 repressed the phosphorylation of signal transducer and activator of transcription 3 (STAT3) both in vivo and in vitro. Activation of the STAT3 signal pathway by Colivelin partly reversed the suppressive effects of rTβ4 on IL-17 expression in vitro.

rTβ4 could alleviate ongoing autoimmune dacryoadenitis in rabbits, probably by suppressing Th17 response via partly affecting the STAT3 pathway. These data may provide a new insight into the therapeutic effect and mechanism of rTβ4 in dry eye associated with Sjögren's syndrome.

Corneal alkali burns are a severe disease and commonly encountered in the emergent clinic. A rapid medical treatment for the burn is very important. Gly-thymosin β₄ (Gly-Tβ₄) is a biomimic derivative of natural thymosin β₄. The aim of this study is to evaluate the corneal recovery effects of Gly-Tβ₄ topical therapy on alkali burns in rabbit corneas.

Rabbit alkali burns were induced with NaOH-contained filter paper. Phosphate-buffered solutions at pH 7.0, Gly-Tβ₄ solutions, blank in situ hydrogels, and Gly-Tβ₄in situ hydrogels were dropped on the burned corneas. The treatments were continued for 14 days. Conjunctiva hyperemia, corneal edema, intraeye extravasation, hemorrhaging, corneal neovascularization (CNV), and corneal opacity were observed. Corneal immunohistochemistry and histopathology were performed.

Gly-Tβ₄ solutions led to a lower corneal burn index than the other regimens. Hydrogels may stimulate the burned corneas due to the direct contact of them, and prevent the rapid release of Gly-Tβ₄. Gly-Tβ₄ significantly inhibited CNV according to the images of the corneas, CNV areas, and CD31 expression. Furthermore, Gly-Tβ₄ improved corneal epidermal recovery according to the histopathological result.

Gly-Tβ₄ solutions are a promising formulation for topical treatment of corneal alkali burns.